Transcript Classification, nomenclature, taxonomy,identification

Classification, nomenclature,
taxonomy,identification
• Classification is arrangement of bacteria into groups
( the same organisms can be classified differently
according to the view: serotype classification,
antimicrobial resistance classification…)
• Nomenclature (name) is the mean of communicating it is binominal
• Taxonomy - science of classification, identification,
nomenclature and making a system
• Identification is practical use of classification criteria to
distinguish certaine organism from others
Graduating
• subspecies: serotyping Streptococcus pneumoniae
type 8
• species - distinct organism with certain
characteristic features, similar organism within
genus Streptococcus pneumoniae, S.sp
• genera:Streptococcus,
• families: Micrococaceae,
• orders
Approaches to taxonomy
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Numerical,
phylogenetic,
genome size,
Guanin+cytosine content,
DNA relatedness, thermal stability of DNA
sequences, DNA relatedness under supraoptimal
conditions…….
• In practice - polyphasic approche - depending on
importnace and purpose of nomenclature
Bacterial identification in
practice
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Pure culture
Colony morphology
gram staining - preliminary identification G+Ggrowth characteristics aerobic, anaerobic
Biochemical properties genus + species
antigenic properties type
binominal nomenclature - clinical purposes
Molecular and genetic characteristic
G+C, DNA homology, thermal stability -epidemiological
and forensic puroses
Principles of diagnosis
• Symptomatic patient - manifestation of
infecton
• Suspection of microbial ethiology exogenous or endogenous
• Specimen selection, collection • Specimen processing and microbiological
examination - specific techniques
• Result communication and consultation
• Asymptomatic patient
Practical taxonomy
Bacteria
Gram positive
lGram negative
G+cocci
Gram+rods
G-cocci
G-rods
anaerobic
aerobic
anaerobic
aerobic
anaerobic
aerobic
aerobic
anaerobic
Peptostreptococcus
Peptococcus
Sarcina
Ruminococcus
Staphylococcus
Micrococcus
Streptococcus
Enterococcus
Clostridium -spore
Bacillus
Corynebacterium
Arcanobacterium
Rhodococcus
Veillonella
Megasphoera
Neisseria
Moraxella
Branhamella
Vibrionaceae
Campylobacter
Helicobacter
Bacteroides
Porphyromonas
Prevotella
Fusobacterium
Gemella
Pediococcus
Leuconostoc
Lacotococcus
Non-spore
Propionibacterium
Mobiluncus
Bifidobacterium
Listeria
Erysipelotrix
Gardnerella
NONFERMENTERS
Pseudomonas
Acinetobacter
Stenotrofomonas......
Leptotricha
Wolinella
Miscellanous bacteria
Mycoplasma, Ureaplasma, Chlamydia
Ricketsiaceae
Spirochetales - Spirochetes, Leptospira, Borrelia
Mycobacterium
Nocardia , Actinomycetes,
ENTEROBACTERIACEA
Bordetella
Pasteurella
Franciscella
Brucella
Haemophilus
Legionella
Bartonella
Eikenella...
Lectures 1,2
• Micrococcaceae (Staphylococcus),
Streptococcaceae G+cocci
• Neisseriaceae: G- cocci
• Bacillaceae, Corynebacterium: G+rods Listeria,
Erysipelotrix,
• Mycobacteria: G- acid fast rods
• Actinomyces, Nocardia: G+rods with special
characteristic
Staphylococcus
• Pathogens of man
• Divided in 2 groups acc to plasmacoagulase production artificial
PC negative - common comensales of human skin and
animals, some of them cause infections in some
circumstances (Staphyoloc. epidermidis, warneri,
haemolyticus, saprophyticus, hyicus)
PC positive - Staphylococcus aureus, St. intermedius
G+cocci in clusters - gram
staining
Cocci in clusters in fluorescein
preparation
Growth characteristic – growing on salt media – staphylococci
Fermenting manitol –yellow – St. aureus vs. St.epidermidis
Staphylococcus aureus G+coccus
• colonizes nasal passage and axillae
• able to grow on salt medium (10%), catalase +, grows in
clusters, PC positive - distinguishing factor
• Structure: capsule, peptidoglycan,protein A, teichooic acid,
clumping factor, cytoplasmic membrane
• Toxins: alfa, beta, delta, gama, leukocidin, exfoliating,toxic
shock syndrome TSST,enterotoxin
• Enzymes: coagulase, catalase, hyaluronidase,
staphylokinase, lipases, fibrinolysin, nuclease, penicilinase
Virulence factor and patogenesis
• Multifactorial
• adherence
surface proteins (att.to fibronectin),
fibrinogen/fibrin binding protein (att. to clots and traumatised
cell, the most important part of slime on biomaterial of
indwelling devices),
fibronectin
• avoidance of host defence
capsule polysaccharide,
protein A - bind IgG - disruption of opsonisation,
leukocidin - toxic for PMNL
• damage of host tissue
- toxins and enzymes
Toxíny
• alfa - cytotoxický pre bb, rozrušuje bunkovú membránu a hladkú
svalovinu ciev, nekrotizujúci
• beta - sfingomyelináza C, termolabilný, hydrolýza fosfolipidov,
deštrukcia tkaniva, tvorba abscesu
• delta - termostabilný, cytolýza, detergentné vlastnosti
• gamma - erytrocytolýza
• leukocidín-zvyšovanie permeability a tvorba pórov v bb. membráne,
odolnosť voči fagocytóze
• exfoliatívny toxín - zodpovedný za SSS – skin scaled sydrome,
rozrušuje medzibunkové spoje - dezmozómy v stratum granulosum
kože, tvorba protektívnych protilátok - len u detí
• TSS-1 - vzniká pri raste u niektorých kmeňov St.aureus TSS – toxic
shock syndrome – toxický šokový syndróm
• enterotoxíny - odolný voči hydrolýze žal. kyselinou, termostabilný, 5
typov A-E, B - pseudomembranózna kolitída, neurotoxín, hnačky a
zvracanie
Enzýmy
• Koaguláza - viazaná (fibrinogén - fibrín) a voľná (cez
medziprodukty), tvorba fibrínovej vrstvy a abscesu - ochrana pred
fagocytózou
• Kataláza - premena toxického H2O2 -- H2O + O2
• Fibrinolyzín- stafylokináza - rozpúšťa fibrínový trombus
• Lipáza - rôzne typy, prežívanie stafylokokov v seboroických
oblastiach - prenikanie do kože a podkožia, tvorba povrchových
kožných infekcií
• Hyaluronidáza - rozrušuje mukopolysacharidy v spojivovom tkanive,
šírenie
• Nukleáza - termostabilný E
• Penicilináza - beta laktamáza, enzým rozrušujúci betalaktámový kruh
PNC antibiotík
Klinické manifestácie infekcie
spôsobenej St. aureus
Hnisavé
• Koža – hnisavé ochorenie kože: vredy, abscesy, akné,
karbunkul, impetigo,
• Systémové - endokarditída, pnuemónia,bakterémia,
osteomyelitída, septická artritída, flebitídy, mastitídy,
meningitída osteomyelitis, infekcia moč. mechúra
• Nozokomiálne infekcie a infekcie kože, katétrov,
popálenín
• GIT infekcie – enterotoxín - zvracanie, hnačky
• TSS - toxic shock syndrom
SSS - scalded skin syndrom
Superantigen: enterotoxins and
TSST
• TSST - exotoxin secreted during the growth of some
strains, connected with superabsorbent tampons
- fever, hypotension, shock, rash, desquamation, multi
organ involvement
TSST-1: 75% menstrual TSS., TSST-2 enterotoxin B and
C: 50% non menstrual TSS
• Enterotoxins A-E- resistant to hydrolysing gastric
enzymes and to heat 100*C (neurotoxin)
• Superantigen: superstimulation of T cells nonspecifically
1 of 5 cells instead of 1 of 10 000 with APC - cytokines
released in large amounts - (any T cell with Vb element is
stimulated)
Clinical manifestation of
St.aureus
• Skin - stye, soil, carbuncule, impetigo, endocarditis,
pnuemonia,
• General pyogenic Nosocomial and wound infection,
infection of indwelling devices, burns,
immunosuppression, phlebitis, mastitis,meningitis
• Toxic GIT infections - emesis, diarhea
TSS - toxic shock syndrome
SSS scalded skin syndrome
osteomyelitis,
impetigo
folikulitis
carbuncul
Stafylococcal scaled skin syndrom
SSSS– Riter´s disease
St. epidermidis and other PC
negative
• Infection of indwelling devices - catheter, shunt,
artificial joint
• Infection of valve - artificial or damaged endocarditis - indolent course - 1 year after surgery
• Staphylococcus saprophyticus - urinary infection of
young sexually active women
• Contamination or ethiological agens
ATB susceptibility and resistance
• Penicillin, staphylococal penicillin - oxacillin,
methicilin,TTC,CMP,ERY, KANA, GEN, STM,
Fluorochinolony
• Quick development of resistance : penicilinase
• Now some strains are resistant to convential ATB
• Hospital strains resistant to many ATB - including
glycopeptides, vancomycin, teicoplanin
• MRSA - meticilin resistant St.aureus
• VRSA - vancomycin resistant St.aureus - resistance
transfered from enterococci
• St.epidermidis - - often meticilin resistant.
Nosocomial problem
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MRSA
multiple resistance
VRSA
resistance to antiseptics and disinfectants
Mechanism of resistance acquiring:
1)extrachromosomal plasmids, transposons
or DNA insertion
2)mutation in chromosomal genes
G+cocci catalase negat.
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Genus:
Streptococcus:
Enterococcus
Aerococcus, Gemella, Lactococcus,
Leuconostoc, Pediococcus - very rarely
pathogenic in men
G+cocci, catalase negat.
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genus
Streptococcus:
Enterococcus
Aerococcus, Gemella,
Lactococcus, Leuconostoc,
Pediococcus
- seldom pathogenic for human
shape
G+cocci in chains or diplococci
Streptococcus - taxonomy and properties
• Taxonomy
- according to haemolysis on blood agar :
alfa - incomplete,
beta - complete,
gama - without haemolysis
- according to serological groupes sc. Lancefield A-H, KV, not all streptococci have this groupe specific antigen of
cell wall
• A
S. pyogenes bacitracin beta hemolysis
• B
S. agalactiae CAMP
beta hemolysis
• C
S. anginosus
beta, alfa
• D
S. bovis
alfa, gama
• S. pneumoniae optochin +
alfa
• S. salivarius
optochin alfa
Streptococcus alfa hemolysis
• S. pneumoniae optochin +
alfa
• S. salivarius
optochin alfa
• - Streptococcus beta hemolyticus
Streptococcus alfa hemolyticus
Streptococcus – beta hemolyticus
A
B
S. pyogenes beta hemolysis (bacitracín+)
S. agalactiae beta hemolysis (CAMP)
Streptococcus sk A - pyogenes
• In bouillon long chains
• Antigenic structure - capsule - hyaluronic acid -identical
with connective tissue - nonimunogennic
• peptidoglycan
• groupe and type specific antigens most important M antigen - on the surface of fimbriae,
virulence
- T protein, R protein, F protein,
• lipoteichoic acid
Factors of patogenity and imunity
• Capsule nonimunogennic - inhibits phagocytosis
• M protein - protection against phagocytosis and
cooperation with C´. Type 1,3,18 - invasive
diseases., M3 a M18 - Rheumatic fever
• F protein - receptor for fibronectin (matrix of
eukaryotic cells - adhesin
• Lipoteichoic acid - ?
Toxins
• Exotoxins - erytrogenic toxin -termolabil, type A,B,C hypersensitivity, endotoxic, cytotoxic, non specific mitogen T and
imunosupresive B lymfocytes activity, rash at scarlatina
• Dick´s, Schultz´s test
• Streptolysin S - oxgen stabile, lysis of ery, releasing of lysosyme,
nonimunogen
• Streptolysin O - reversibily inactivated by oxygen, imunogen,
antibodies against streptolysine O - ASLO, killing of leukocytes
• Streptokinase - lysis of blood trombus, spread of bacteria
• DN-ase - noncytolytic, depolymerisation of free DNA - in pus,
declines viscosity of pus, spread
Streptococcal infections
• Streptococcus pyogenes HSA
- pharyngitis - dif dg. from viral
- scarlat fever - pharyngitis with exanthem - infectious strain
gained by lysogenic conversion ability to produce
erytrogenic toxin (rash with desquamation of skin,
circumoral whiteness, strawberry thongue)
- toxic shock syndrome - cellulitis, necrotising fasciitis,
hypotension, erytrodermia, multiorgan failure, bakteraemia,
M1, M3, M18 - types
- erysipelas, pyodermia - purulent skin diseases
Flesh eating streptococcus
Toxic shock syndrome
– celulitís and necrotising faciitis - superantigen
Streptococcal infections- late sequelae
• Febris rheumatica - rheumatic fever - nonpurulent
streptococcal disease - inflamatory disease of heart, joints,
vessels and submucosis - autoimmune disease related to
some serotypes of M protein ( types specific antigen, factor
of patogenity, numbered M18, M3…) sequelae present
only after respiratory infections. Not after skin infection
• Poststreptococcal acute glomerulonefritis - specific
nefritogen strains, after skin and respiratory infections
Diagnosis - laboratory
• Microscopy - gram staine, G+ cocci in chains, not
colonising skin, in skin swab together with leu significant for disease. Of any value from URT sample
• Detection of antigens - directly from clinical material
(also from URT), detection of groupe specific antigens of
cell wall, specific, not sensitive - negative tests should be
confirmed by cultivation
• Cultivation, Identification - hemolysis, bacitracin test
• Antibodies - ASLO - confirmation of preliminary
streptococcal infection in patients with RF a GNF (+ anti
Dnase antibodies
Streptococcus agalactiae, HSB
• newborne infections (menginitis, pneumonia), puerperal postpartum sepsis /colonisation of URT, GIT and vagina danger of contamination of newborne during prolonged and
preterm labour, importance of maternal imunity/
• G+ cocci in chains, beta hemolysis, dif dg.from HSA CAMP test - strenthened hemolysis of Staf. aureus papillon
• Strukture - polysaccharid capsule, cell wall - peptidoglycan
with type and groupe specific antigens and lipoteichoic acid,
cell membran
• Antigenic strukture - antibody against capsule antigens are
protective - diseases in newborne
• Enzymes - Dnase, hyase,protease, hemolysin
Other streptococci
• Beta hemolytical group C,F,G - URT and skin
infections never late complications
• Viridant -alfa a nonhemolytical streptococci
- Streptococcus salivarius, viridans - bacteremia,
subacute endokarditis, caries, intraabdominal
purulent infections
• Important condition for development of disease is
the preliminary damage of tissu ( tooth, valve)
• Carries - formation of dextranu from glucose
Streptococcus pneumoniae
• G+cocci in pairs, diplococcus, lancet shape - candle and
flame,viridant - dif.dg - positivity of optochin test, bile
solubility test - colonies are dissolved by powdre bile
• encapsulated and nonencapsulated strains- polysaccharide
capsule - factor of virulence - more than 90 serotypes based
on capsule antigens
peptidoglycan of cell wall typical for G+ cocci (N
acetylmuramic and N acetyl glukosamin net), teichoic acid
and cholin ( unique substance, important for cell division)
• 2 forms of teichoicacid in cell wall - surface (C substance)
and covalently bound
STREPTOCOCCUS PNEUMONIAE
Streptococcus pneumoniae:
Gram-positíve diplococcus
0.5-1.25 / um
STRUCTURE
G+ diplococcus in CSF
fluorescent microscopy
Laboratry
diagnosis
• CSM –- G-diplococci, leucocytes, antigen detection
- latex agglutination in CSM, and urine
Cultivation of CSM and blood - hemocultivation –
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• ATB susceptibility
OCHORENIA SPOSOBENÉ
STREPTOCOCCUS PNEUMONIAE
 Streptococcus pneumoniae diseases
Non encapsulate
Localised
encapsulated
Invasive
Neinvazívne
Invazívne
- Sínusitída
- Bakteriémia
- Otitis media
meningitída
- Pneumónia
endokarditída
peritonotída
septická artritída
iné (mäkké tkanivá)
CRP
• CRP - C substance of str. pneumoniae During bacterial infecions organism
syntetises CRP - C reactive protein - protein
of acute phase of inflammation, reacting
with C substance of Str. pneumoniae
Streptococcus pneumoniae infections
• Encapsulated strains - virulent - pneumo, (lobal, crupous without ATB resolution after 14 days - crisis - production of
anticapsular antibodies) meningitis, bakteremia - age
predilection -in seniors (superinfection od viral infection of
RT) and children up to 3 years
( imunologically defect, encapsulated strains T independent
- protectiveantibodies ( 90 serotypes, vaccination,
conjugated vaccines)
• Nonencapsulated strains - sinusitis, otitis media
KOLONIZÁCIA A PRENOS
INFEKČNÉHO AGENS
nosičstvo sa môže vyskytnúť až u 60% detí
predškolského veku a 30% dospelých
Asymptomatický
nosič
Aerosol
Inhalácia
Pacient s
pneumokovo
u infekciou
Nosová
dutina
Nasofarynx
miesto
kolonizácie
Trachea
Diseminácia
Invázne
Opúzdrené kmene
!
PATOGENÉZA
Meningitis
!
Sinusitis
Neinvázne
Neopúzdrené
kmene
Otitis media
Nazofarynx
!
kolonizácia
Bakterémia
Artritis
Peritonitis
!
Pneumónia
ATB susceptibility
• Str. pyogenes - 100% susceptible for PNC, (in allergies
macrolides ERY)
• Str. agalactiae - good susceptibility for PNC, some strains
tolerate - inhibition, not resistence, resistence to ERY, TTC
• Str. salivarius - susceptible for PNC,
• Str. pneumoniae - PNC, TTC, CMP, CEF., resistence for
PNC - multiresistence - decline of affinity of ATB to PBP
Enterococcus
• G+ cocci, colonisisng in great quantities intestine and colon,
able to grow in the presence of bile - dif. dg. - esculin media
• E. faecalis
• E. faecium
• urine tract infections, intraabdominal abscesses,
bakteraemia
• ATB nonbactericidal for enterococcie, resistence - synergic
therapia: aminoglykosides + cell wall acting
• Resistence plasmid transferable - (on staphylococci too vancomycine)
G – Coccus
Praktical taxonomy
Bacteria
Gram positive
lGram negative
G+cocci
Gram+rods
G-cocci
G-rods
anaerobic
aerobic
anaerobic
aerobic
anaerobic
aerobic
aerobic
anaerobic
Peptostreptococcus
Peptococcus
Sarcina
Ruminococcus
Staphylococcus
Micrococcus
Streptococcus
Enterococcus
Clostridium -spore
Bacillus
Corynebacterium
Arcanobacterium
Rhodococcus
Veillonella
Megasphoera
Neisseria
Moraxella
Branhamella
Vibrionaceae
Campylobacter
Helicobacter
Bacteroides
Porphyromonas
Prevotella
Fusobacterium
Gemella
Pediococcus
Leuconostoc
Lacotococcus
Non-spore
Propionibacterium
Mobiluncus
Bifidobacterium
Listeria
Erysipelotrix
Gardnerella
NONFERMENTERS
Pseudomonas
Acinetobacter
Stenotrofomonas......
Leptotricha
Wolinella
Miscellaneous
Mycoplasma, Ureaplasma, Chlamydia
Ricketsiaceae
Spirochetales - Spirochetes, Leptospira, Borrelia
Mycobacterium
Nocardia , Actinomycetes,
ENTEROBACTERIACEA
Bordetella
Pasteurella
Franciscella
Brucella
Haemophilus
Legionella
Bartonella
Eikenella...
Family Neisseriacae
Family Moraxellaceae - genera Branhamella,
Moraxella, Acinetobacter
• Genus Neisseria
• Species: N. meningitidis -colonisation of URT or
menigitis, setpicaemia- meningococcaemia fulminant sepsis, pneumonia, arthritis
N.gonorrhoeae - uretritis, cervicitis, salpingitis,
proctitis, septicaemia, arthritis, conjunctivitis,
pharyngitis, Pelvic inflamatory disease
other neisseria - colonisation of mucous
membranes and skin
Colonisation
Asymptomatic
carrier
Aerosol
Inhalation
Pacient with
the infection
Nasofarynx
colonisation
Encapsulated
strains
Colonisation !
Invasion throught
mucous membrane
PATOGENESIS
Meningitis
Non
encapsulated
strains
Nasopharynx
colonisation
Bakterémia
Meningococcaemia
Neisseria meningitis
• Encapsulated (polysaccharide
capsule), G- diplococcus,
coffee beans,
• Serogroups (13)
- A - (1serotype),
- B,C(several serotypes 20),
-Y, W135…
most frequent,
8 immunotypes
Neisseria meningitis
• Pathogenicity:
colonisation of nasopharynx (pilli) - attachment to
noncilliated cell of columnar epitel
spread - escape to phagocytosis - antiphagocytic property
of capsule
toxic effect - endotoxin LPS - vascular damage
(endothelial damage, inflamation of vessel wall, DIC,
trombosis) - hyperproduction of membrane fragments in
growing cells
Neisseria meningitis - clinical
syndromes
• Meningitis - high mortality when not treated 100%
• Meningococcemia - life-threatening,, trombosis of small
blood vessels, multiorgan invovement, petechiae,
overwhelming disseminated intravascular coagulation with
shock - destruction of adrenal glands- Waterhouse
Friderichsen sy, - chronic septicaemia
Neisseria meningitis -therapy
• Therapy - PNC(genetic alteration of PBP ),CMP,
cephalosporin,
• Eradication - (spread of strains) sulphonaminds, rifampicin
• Vaccination - A,C,Y,W vaccine - not for children under 2,
conjugate vaccine - immunogenic for children, B vaccine polysaccharide is weak immunogene
Neisseria meningitis - immunity
• Bactericidal antibodies(serogroup) (only in type B serotype
ab are also protective) and complement - protective
• polysaccharide capsule - susceptibility of children under 3
years - T independent Ag
• susceptibility of infant after decline of maternal antibodies
• Acquired immunity - 2 weeks after asymptomatic
colonisation
• Transmission by droplets, prolonged close contact
(millitary, crowded communities)
• Sporadic and epidemic spread
Laboratory diagnosis
• CSF - Gram stain - G-diplococci, leucocytes,
antigen detection - latex agglutination in CSF, urine
• cultivation of CSF and blood culture - chocolatised
agar + 5%CO2, !not refrigerator! biochemical
properties - dif dg. from other meisseria serogroupe
typisation - agglutination,
• ATB susceptibility test
• from nasopharyngeal swab - dif. dg. from non
pathogenic neisseria - biochemical test
Microscopy .Gram
Latex aglutination
Ag
Cultivation
PCR
Sampling,
transport
Thay Martin
Room
temperature
Cultivation
Chocolate
agar
Oxidase +
Neisseria gonorrhoeae
• Most common sexually transmitted disease (STD)
uretritis, cervicitis, arthritis, conjunctivitis, local
complications
• G- diplococci, 5 different types of colonies (T1-5),
cytochromoxidase positivity, glucose fermentation
• Structure: capsule, pilli - in virulent strains(T1,2) attachment, OMP - outer membrane proteinI.,II.,III, Iron binding protein - removal of iron from
host cells - iron essential for metabolism of
gonococci, LPS-endotoxin, protease, penicillinase
Pathogenesis and immunity
• Attachment - pilli, penetration and multiplication
and pssage throughthe cells into subepitelial space.
Nonpilliated are avirulent. Protein I interfere with
neutrophil degranulation. Endotoxin - tissue
destruction.
• IgG and C´: individuals with C´deficiencis are at
increased risk for systemic disease.
• Only humans, asymptomatic reservoir
• Multiple infections - lack of protective immunity antibodies against pilin protein - antigenic diversity
Clinical signs
• Infection in men - uretritis+complication
• in women- cervicitis - ascending genital inf.
• Disseminated infection with septicaemia, skin, joints
infections:(fever, artralgias-migratory,suppurative
monoarthritis-wrist, knee, ankles, rash over
extremities
• Perihepatitis, purulent conjunctivities in newborne,
anorectal gonorrhoe in homosexual, pharyngitis
• Chemoprophylaxis - - ineffective
Cultivation +ATB
• Neisseria gonorrhoe - PNC – penicilinase production
- chromosome type of resistence - changes in cell surface,
- ceftriaxon. TTC, chinolons, makrolides - azitromycin –
therapy of chlamydia infection
• Microscopy: Vagina swab in susp. gonorrhoe: - Gram staining: G
- diplococci, coffee beans, epitelial cells., leukocytes. From culture
colonies: G- diplococci
• Cultivation: Swab from vagina or discharge – on blood agar,
modified blood agar, chocolate agar + ATB – inhibition of
contaminating flora, Grey cololnies after application of
cytocromoxidase – become black – slide sc. Gram, - biochemical
tests for diff.dg. from other Neisseria
Genus:Bacillus - G+sporeforming bacilli, aerobic
and facultatively anaerobic
•
Bacillus anthracis,Bacillus cereus,Other Bacillus sp.
• Bacillus cereus - 2 enterotoxins
- heat stable - emetic form - contaminated rice - heat resistant spores
survive initial cooking that kills vegetative cells, germinate, multiply
and toxin is not destroyed by reheting
- heat labile - diarrheal form - adenylcyclase-cAMP system
stimulation in intestinal cells - fluid accumulation -contaminated meat
and vegetable - toxin is produced in situ, longer incubaion
• Panophthalmitis - traumatic eye infection (soil, penetrating object),
complete loss, massive destruction
- toxins - necrotic - heat labile enterotoxin
-cereolysin -hemolysin
- phospholipase C - lecithinase
• Ubiquitous, isolation witout symptoms = contamination
• Other Bacillus - immunosupressed patients - shunt and catheter inf.
Bacillus anthracis
• Spore and capsule not seen in clinical specimens
• Antrax toxin - 3 antigenically distinct components: -protective Ag,
lethal factor, edema factor - alone not active,
• Pathogenesis: capsule - antiphagocytic, Ab are not protective, toxins protective + lethal or edema f.
• Human diseases - cutaneous - inoculation - painless papule, ulcer,
necrosis
- inhalation - rapid progresive diffuse pulmonary involvement respiratory failure - 95% mortality
- gastrointestinal - ingestion - rare - mesenteric adenopathy,
hemorrhage, ascites, 90% mortality
• Cultivation - on nonselective media, rapidly growing adherent
colonies, no hemolysis, Non motility,liquifaction of agar
Microscopy: caput medusae - serpentine chains,
• Therapy - PNC (TTC, CMP)., control of animal antrax, vaccination.
Non sporeforming G+ bacilli heterogenous group
• Corynebacterium - coryneforms, diphtheroids- C. diphteriae
(diphtheria), C. jeikeium (oportunistic). C.
urealyticum(urinary tranct inf.), C. pseudodiphthericum
(endocarditis), C. ulcerans (pharyngitis)
• Arcanobacterium haemolyticum - pharyngitis
• Actinomyces - granulomatous ulcerative inf.
• Rhodococcus - suppurative pneumonia, opportunistic
• Listeria - meningitis, septicemia, granulomatosis
infantiseptica
• Erysipelothrix - erysipeloid, septicemia, endocarditis
Corynebacterium
• Pleiomorphic G+ rod, arranging to short chains,
forming china letters
• Metachromatic granules – the final colour is
defferent from original stain and from other parts
of cell
• Special cell wall structure – mesodiaminopimel
acid, mycolic acid – taxonomically near
Mycobacteria
• C. diphtheriae – preventable disease – inhalation –
asymptomatic carriage or disease
• C. JL – jeikeium – opportunistic pathogen
hematological disorders
Pathogenesis
• Diphtheric toxin – tox gen – lysogenic
conversion
• AB toxin – blacking protosynthesis –
prolongation of peptid chain on ribosomes
• Schick skin test – detection of neutralisation
antibodies when i-d appôicated diphtheria
toxin
Lysogenic conversion
• Strain of C. diphtheriae gains his
toxigenicity by lysogenic conversion thanks to bacterophage. Bacterial virus bactrophage is able to transfere genetic
information about the production of toxin tox gen - fom one cell ( toxigenic) to the
other (non toxigenic) C. diphtheriae cell.
Enzymes
• Phospholipasa D – dermonecrotic toxin –
spread – increasing of vascular permeability
– C. ulcerans
• Urease – alkalinisation of urine calculi C.
urealyticum
• ATB resistance – selection of resistent
strains – C. urealyticum, C.JK – resistent to
ATB used for urinary infections
Clinical syndromes
• Diphtheria – depend on immunity and place of
infection – asymptomatic colonisastion. Mild
infection or fulminant diphtheria
• Pseudomembrane on tonsils, pharynx, nose and
general symptomes and complication
(myocarditis)
• Skin diphtheria
• Therapy –ATB .PNC, ERY – elimination of
bacteria, not threating intoxication, antitoxin,
immunisation
• Debacilisation of carrierers –
• Protective immunity in skin forme
Lab. dg. of corynebacterium
• Diagnosis of the disease is based on clinical picture and
epidemiological history. Lab. dg. is long and complicated
Appropriate sampling
• Microscopy - smear with Gram staining and Albert´s staining does
not distinguish between C. diphtheriae and other Corynebacteria.
• Cultivation: Blood agar - 3 typs od colonies - mitis, gravis,
intermedius, Loffler´s medium, tellurit medium brown colonies with
halo
• Biochemical identification- differentiation from other corynebacteria
present in throat
• toxigenicity of the strain - ELEK,
• ATB susceptibility
Cultivation of Corynebacteria
• C. diphtheriae - blood agar
• C. pseudodiphthericum - blood agar, inverse
CAMP
• C. diphtheriae on tellurit medium - brown
colonies with brouwn ring - halo
Detection of toxigenicity of C.
diphtheriae
• ELEK´s test of toxigenicity - imunodiffusion of
suspension of tested strain and antidiphtheric serum
in agar - zone of precipitation
• Annimal mode: application of
diphtheria toxin i.d. ………………..necrosis
• antidiphtheric serum (i.p. or i.d.) + toxin …no
necrosis
Arcanobacterium
• A.haemolyticum - colonise human, responsible for
pharyngitis (+/- scarlet fever-like rash), cutaneusinf.,
endocarditis, meningitis - in older patients
• Missdiagnosed as Str. pyogenens, grows slowly,
weak hemolysis
• 2 toxins - hemolysin and phospholipase D
• enzymes neuraminidase
Erysipelotrix
• E. rusiopathiae: G+ non spore forming facultative anaerobic
bacillus, worldwide distribution in animals
• Cultivation in reduced oxygen athmosphere, small grayish,
alfa hemolytic colonies, 2-3 days, sample from deep tissue
or deep aspirates.
• Erysipeloid - occupational disease (butchers, meat
processors) after subcutaneous innoculatio -localised skin
infection
-generalised cutaneous forme
-septicaemia - associated with endocarditis (undamaged
heart valve - aortic)
• Therapy PNC, ERY,CLI (sulfonamids, vancomycin, aminoglycosides resistent)
Listeria
Rhodococcus
• G+, obligate aerobic, red-pigmented, acid fast,
mycolic acid. Veterinary pathogen. Present in soil
• Intracellular - surviving in macrophages
• Granulomatous inflammation with abscess
formation (lung,lymph nodes, menings, pericardium,
skin) - immnosupressed
• Cultivation - nonselected media, aerobically,
pigmented colonies after 4 and more days
• Therapy : prolonged - multiple ATB able to
penetrate into macrophages
Gardnerella
• Morphologically resemples gram neganive bacilli, has cell
wall structure of gram +, nonmotile, not capsule
• part of normal vaginal flora
• present in bacterial vaginosis together with obligate
anaerobes - Mobiluncus, Peptostreptococcus,absence of
Lactobacillus. Present in postpartum bacteremia,
endometritis, vaginal abscesses
• Lab. dg. - simple isolation is not prooving, importance of
microscopic examination - clue cells - epitelial cells covered
with G variable bacilli (Gardnerella) and G- small curved
bacilli (Mobiluncus), absence of G+ bacilli (Lactobacilli)
• Therapy: ampicilin, metronidasol
Nocardia
• G+strictly aerobe rods. Similar to quickly growing mycobacteria,
saprophytes in environment. Acid fast., Mycolic acid
• Nocardia asteroides, N. brasiliensis, N. madurae
• pneumonia - with confluent abscess formation, exogenous
inhalation
• skin infection - A. brasilinensis - localised celulitis, purulent sinuses
with chronical granulomatous inflamation - mycetoms
• Madurmycosis - chronical granulomatous infection of bone and soft
tissue, deformations, (Sudam, Northern Africa, East India)
• diseminated - CNS - brain abscesses, in immunocompromised
• Lab. Dg. - microscopy - modified Ziehl Neelsen, Gram +,
cultivation - standard media -2-30 days, colonies adherent to agar,
cream, orange rose color, chalky consistence
• Therapy: surgery+ATB 3 months sulphonamids, amikacin,
imipenem, broad spectrum to be effective if fungal ethiology
Lectures 3,4,
•
•
•
•
•
Anaerobes
Enterobacteriacea
Vibrionaceae
Campylobacter
Helicobacter
Anaerobe gram + sporulating rods
• Clostridia - 100 species, some are aerotolerant - C. histolyticum
some looks lide G-. Commonly present in soil, water, GIT
• C. perfringens - Bacteraemia, myonecrosis - gas gangren, infection
of soft tissu, necrotising enteritis
• C. tetani - tetanus
• C. botulinum - botulismus - infant, wound, food intoxication.
• C. difficile - pseudomembranous colitis connected with ATB, 2
toxins, part of FMF, exposition to ATB kills other colon flora overgrowth of C. difficile
• C. septicum - not injury connected myonecrosis, patients
imunodepresed - Ca of colon - interruption of colon wall integrity
and spread of Clostridium to tissues
Clostridium tetani
• Motile rod, sporulating, G+ often looks like G-. Very sensitive to
oxygen, metabolically poorly active.
• Heat labile neurotoxin- tetanospasmin - released after lysis of cell.
AB toxin - block of neurotransmmiters in CNS on inhibitory
synapses - not regulated excitationon synapses - spastic paralysis
• present in soil, GiT. Vegetative forms are sensitive to oxygen.
Spores survive for years
• Generalised tetanus, localised tetanus, tetanus of new bornes - drug
abusers tetanus
• Therapy - PNC, antitoxin, - toxin bound on nerve endings is
protected and cannot be neutralised. Symptomatic therapy
• Vaccination
• Lab dg. - toxin detection - annimal model, microcopy - G+rod with
spores terminally located - enlarging the rod
Clostridium botulinum
• Heterogenous group of nutritionally requiring sporulating bacteria 4
groups I - IV based on proteolytical activities and type of toxin. 7
antigenically different botulo toxins A,B,C alfa, D, E, F, G - A,B,E
most frquent. AB toxin , heat labile - 20minút /80*C
• blocking neuromuscular transmition on synapies of periferal nerves.
• After consumption of contaminated food, cans., wound botulismus or
infant botulismus after honey consumption in infant less than 1 year.
• Blurred vission, dilated pupils, dry tongue, constipation, aches, flacid
palsy - complete recovery after years.
Wound botulismus - very rare
Children botulismus- 1976 - in vivo production of neurotoxin from
colon colonisating clostridia (in 6m - 1y of age) - progresive flacid
palsy, - mortality 1% - often attributed to other reasons( SIDS)
• Dg. - clinical, detection of clostridia and toxin
Th. - antitoxin, PNC- destruction of spores and prevention of
germinating of spores
•
Clostridium
perfringens
Colonisation of serious disease, hemolysis, biochemical activity.
Production of letal toxins (alfa, beta, epsilon, iota, heatlabile
enterotoxin) with life threating biological effect and ensymes. Types
A-E: A v in environment, B-E in colon.
A gas gangrene and intoxikcation, C necrotising enteritis
• Bacteraemia - often not significant, transient from contaminant from
skin.
Gas gangrene - life threating, histotoxic clostridium, after injury, very
painful, devitalised tissue with gas production. (C. septicum, histolyticum,
novyi)
Celullitis, fasciitis - after wound colonisation - often not significant or very progresive destruction ( C. septicum)
Necrotising enteritis - intestine, type C, 50% lethal
Intoxiction from food - short incubation time, watery diarhea ingestion of contaminated food, toxin - heat labile protein
• Microscopy - G+rod withou leu from clinical material
Th - surgery, high dose of PNC, antitoxin, hyperbaric chamber
G+rods anaerobes
• Not sporulating
- Actinomycetaceae Actionomyces israeli, A.naeslundii
---cerebral, cerevicofacial, abdominal actinomycosis, chronic purulent disease
with abscess formation. Lab.dg. Samplling from depth of sinuses, sulphur
granules, colonies in shape of tooth - molar, requiring and long cultivation,
Th: surgery +ATB prolonged PNC
-Propionibacteriaceae Propionibacterium acnes
--- colonisation of skin, external ear, conjunctive. Opportunistic infection in
patients with foreign bodies. Lab. dg. On common media - anaerobe
-Mobiluncus ---gram variable, morphology of G+,
requiring cultivation,
colonisation of genital tract of women - vaginosis - defect of eubiosis,
overgrowth - not inflamatory disease
-Bifidobacteriaceae, Eubacteriaceae,- Lactobacillaceae . -part of
FMF of
GIT, femal genitals
• Sporulating - strictly anaerobe, spores, tykpicall situated,
proteolytical
G-rods anaerobe
• Bacteroides fragilis - obligate anaerobe rods, not sporulating, important
part of FMF in oropharynx, UGT, GIT. - pleuropneumonia, intraabdominal
abscess, genital infection. Endogenous a polymicrobe infections, formation of
abscess in destruction of barieres. Lab.dg. strict anaerobe sampling and
cultivation and transport , stimulation of growth in 20% of bile. Th surgery +
ATB, - production of betalactamase, Metronidazol
• B. stearothermophilus- sporesy used for testing of effectivity of sterilisers
• Prevotella, Porphyromonas, Fusobacterium, Leptotricha,
Wolinella
G+cocci anaerobe
• Peptostreptococcus, -Peptococcus,- Sarcina,
- Coprococcus, - Ruminococcus
-colonising skin and mucous membrane of GIT, UGT, oportunistic
pathogens often in connection with foreign bodies, requiring
cultivation, slowly growing
- pleuropneumonia after aspiration, sinusitis, brain abscess, spread
from orofarynx or lungs, intraabdominal infection and sepsis spread
from colon, pelvic infection (endometriis, abscess, sepsis, vaginosis)
infection of soft tissue (celulitis), endocarditis, osteomyelitis
• Lab.dg - differenciation from colonnising flora, transport and
sampling under anaerobe condition, prolonged cultivation
• Th - PNC, cefalosporins, imipenem, CMP - often polymicrobial
ethiology.
• G-koky anaeróbne
• Veillonella, Megasphera -present in orofarynx, low
virulence, in cultivation and diseases usually present
in mixture
Physiology and structure of
Enterobacteriaceae
• G- rods, usually motile (flagella, not spore forming, facultative
anaerobes, nutritionally not requiring, biochemical active
catalase +, oxidase -, cytochromoxidase COX - = dif.dg.from
Pseudomonadaceae
• Lactose fermentation + or -, rezistence to bile salts, capsule - dif.dg
from other Enterobacteriaceae
• 3 types of antigens:
• - somatic O antigen - most important cell wall ag, heatstabile, LPS
lipopoly saccharide consists of 3 parts - O polysaccharide, core
polysaccharide and lipid A - with endotoxin activity
• - capsular K antige - heatlabile cross reacting with antibodies against
other bacteria. In Salmonella typhi under name of Vi antigen
• - flagellar H antigen - heatlabile, antigen and phase variation
Biological effect of endotoxin
• Part of G- bacteria cell wall - released after its destruction by ATB
• Fever
• leukopaenia followed by leukocytosis
• activation of complement
• Thrombocytopaenia
• Disseminated intravascular coagulopathy DIC
• decreased periferal circulation and perfussion of big organs
• shock
• Present in GIT
E. coli
• causes - endogennous infections after breaking of immune barieres from
physiological flora ( z focus in UGT or GIT, most important cause of G- sepses),
Infection of urinary tract (most out patients infections - rises from GIT strains,
specific serotypes binding with specific adhesis to epitel receptors),
- gastroenteritis
- neonatal meningitis
• enterotoxigen ETEC - production of heatstabile and heat labile toxin, mild water
diarhoea, travellers diarhoea,
• enteroinvsive EIEC - spread to epithelial cell of the colon, destruction of epithel,
fever, blood and leu in stool, Confirmation of invasivness - Sereny´s test innoculation of strain do conjunctiva of annimal- results in inflamation
enteropathogenic EPEC - serious child diarhoea , shiga-like toxin, able to adhere
to erytrocytes
• enterohemorhagi EHEC - production of verotoxin. From non complicated
diarhoea to hemorhagic colitis and . Hemolytic-uremic sy - type 0157:H7, - acute
kidney insufficiency, trombocytopaenia, in children under do 5 r.
Salmonella
• 1500 serotypes, different names according to place of identification
• DNA analysis = 1 species (S. enterica) and its 7 subgroupes
• Present in annimals, spread by contaminnated food - primary from
contaminated annimal products - eggs - salmonella disease of hens.
Secondary - from carrier or patients. Need of big innoculun 106-8 spread by food where it can multiply
• Salmonellosis - enteritis - infection of colon, fever, nausea, vomiting,
headaches. Dg.: Stool sample 3 x consecutively. Lactose negative
colonies, biochemical identifiction. Serotyping by slide agglutination
according to Kauffmann White scheme - epidemiological purposes
(S. enteritidis, S. infantis, S. agona, S. kentucky…………………….)
Salmonella typhi
• Typhoid fevers - fever with deterioration - systemic
disease - penetration to lamina propria, lymphatic nodes,
RES (multiplication in hepari - spread via bile vesicle to
intestine - , spleen, bone marrow), blood - bakteraemia,
exanthem - roseola, fever, headache. GIT symptomes not
always present - perforation . Carriage of Salmonella.
• Dg. - sample of stool, repeated after stimulation by
MgSO4. Typical lood of cultivation. Serotypisation by slide
agglutination. Detection of antibodies Vidal reaction. Vi
agglutintiona
• Th. - CMP
• Salmonella paratyphi A, B, C - Typhus like, milder
Shigella
• 4 species - Sh. dysenteriae, Sh. flexneri, Sh. boydii, Sh.
sonnei a 38 serotypes
• Dysentheria - watery diarhoea with blood, often
epidemic, transmitted with contaminated hands.
Surviving in water 6 months. Small infectious dosis 200 bacteria. Disease of small intestine first withou
invasion. Production of enterotoxin - invasion and
destruction of epitel, ulcers formation - pus and blood
in stool
• Dg. Based on biochemical activity - characteristics of
cultivation - without smell, not gas, lactose negative.
Serotype differenciation - slide agglutination
Yersinia
• 7 species - Y.pestis, Y. pseudotuberculosis, Y. enterocolitica, +
oportunistic Yersíniae
• Y. pestis - plague - urban and forest type, not GIT disease.
Adapted to i.c. parasitismus, not surviving in nature. Virulence
factors - i.c. surviving, , polysaccharide capsule, endotoxin, .
Urban plague - circulating between rats, transmitted by insect to
man during rodent bacteraemia. Replication in colon of insect and
transmission to other rodent or man is on by chance.
Forest pôague - Not controlled.
• 2 clinical formes - bubonic - 7 days after biting by insect - painful
lymphadenopathy75% lethality
- pneumonic - 2 days after inhalation - fever, pneumonia,
inhalation spread by droptets, epidemia letality 90%
Yersinia enterocolitica
• Enterocolitis in colder environment, most
activity in 22*C, food borne infection,
diarrhoea, fever. Chronic disease - terminal
ileum, mesenteritis immitation of appendicitis.
Extraintestinal symptomes - arthritis, hepatitis,
osteomyelitis • Dg. - cultivation or serology by agglutination in
extraintestinal sy
• blood borne infection - bacteraemia and
endotoxic shock (blood cans - cooled)
Klebsiella
• G- rod with typical structure
• Encapsulated– mucous look of colonies - increased
virulence, notmotile
• Klebsiella pneumoniae – pneumonia in patients not able
to clean from bronchial stroma and lungs the aspirate
Necrotic destrukction of alveols. Bloody sputum.
Infection of tissue and urinary tract
• K. rhinoscleromatis - sclerom
• K. ozaenae – atrofic disease of nose musous membrane
Vibrionaceae
• G- curved rods aerobe and anaerobe growth. Present in
water
• Oxidase test – posit. – diff. Dg from Enterobakteriaceae
• V.cholerae - gastroenteritis
• V. parahaemolyticus - gastroenteritis
• V. vulnificans – bakteraemia, infection of wound,
celullitis – exposition to contaminated water, GIT – row
sea fruits
• V. alginolyticus – infection of wound and soft tissue,
otitis externa
• Other vibrias – infection of wound and moderate GIT infections
• Plesiomonas – GIT infections from raw sea fruits
• Aeromonas – opportunistic (…pijavice, after microsurgery)
V. cholerae
• Growing in 18* - 37*C
• Serologically – 6 groups based on O antigen (patogennic are O1)
• Vibrio cholerae O1 – 2 biotypes – el tor a cholerae and each present in
2 serologic subgroupes – ogawa a inaba
• Adherence, not invasive.
• Production of AB toxin – cholera enterotoxin – binding on specific
receptors of intestin, causing secretion of Na, K a bicarbonate to
lumen – liquid lost - 1 liter in hr. (hypovolaemia, arythmia, kidney
insufficiency). Not adhering bacteria are avirulentné
• In waters of Asia, human carriage and sea annimals. Spread by
contaminated water and food. Necessity of big innoculum – 103-5.
Less in condition of patients achlorhydria ( 104 bacteria after
bicarbonate)
Diagnosis and therapy of Vibrio
• Microscopy – small, motile – native smear or
darkfield
• Cultivation – selective media – not supporting
acid environment and drying– enriche alkalin
buillon pH 8,6
• Dif. dg. V. cholerae a other vibiras
(halofile, requiring 1%NaCl)
• Therapia V. cholerae – ATB for debacilisation
only, exotoxinu. (TTC,ERY, CMP, COT).,
symptomatic - solution replacement V.
parahemolyticus – mild disease, would infection –
ATB – TTC)
Campylobacter
Comma shape, G-, motile, 11 species,7 subspecies
Requiring microaerophil defined environment, 42*C,
passing through bacterial filters - selective isoloation
Campylobacter jejuni – gastroenteritis in patients with
decreased immunity –hypogamaglobulinemia, decreased
acidity of stomac liquid. Infection of epitel of jejunum,
ileum and colon results in oedematous , bloody mucus
with abscess formation.Infiltration to lamina propria.
Enterotoxin, cytopatic toxin, endotoxin. Poultry, food
neutralising pH reduced infection dosis.
C.coli - gastroenteritis, C. fetus – septicaemia, artritis,
trombophlebitis, meningitis
Lab dg.: cultivation on selective media, microscopy - of
stool - leu, antigen detection, Th.: ERY
Helicobacter
• Spiral rods, G-, in connection with gastritis, gastric and
duodenal ulcer, susp.Ca – Gastrospirillum hominis
• Pathogenesis – production of urease – formation of
cloud of NH4, that protects bacteria from acid in
stomac - motility - mucinase formation – spread to
mucous layer - adherence – attachment to intracelular
junction – inflamation with mononuclear infiltration in
lamina propria.Escape to IgA specific antibodies
because of its location in mucous layer
• Interhuman transmission
• Th. ATB . Bismuth, nitroimidasol a amoxicilin or TTC
Laboratory diagnosis
• Biopsy taken during endoscopy: Microscopy –
hematoxylin eosin staining, Gram staaining. !
Desinfection and contamination of endoscopic
devices
• Detection of alkalic metabolits of urease activity.
Rapid testt – 1-2 hrs – from bioptic material or
breath test
• Cultivation - from bioptic material – enrichee
media – blood, haemin, charcoal - problems:
activity of inhibition caused by desinfection of
endoscopes
• Serology: Detection of IgA ! Interpretation –
prolonged presence not discriminating between
acute and pass infection.